Approximately one-third of the human proteome is comprised of membrane proteins that belong to protein families with a variety of biochemical activities, such as transporters, channels, receptors, recognition molecules, and adhesion molecules. Membrane proteins are critically important molecules for cell survival, maintenance of cell homeostasis, cell signaling, immune surveillance, molecular transport, and cell-cell communication. This class of proteins represents up to 70% of therapeutic targets for all prescribed drugs. Therefore, development of a high-throughput platform that enables profiling membrane proteins in an active conformation for their biochemical activities would have an important impact on drug discovery by streamlining small molecule screening methods. However, membrane proteins, especially those carrying multi-pass transmembrane (TM) domains, are notoriously difficult to study because they have to be embedded in a membrane to maintain a native conformation and many require proper posttranslational modifications (PTMs), such as glycosylation, which occurs during transport in the cellular secretory pathway. Although membrane protein microarrays have been reported previously, biochemical purification using detergents limits the throughput and subsequent manipulation. (Fang et al. (2002) J. Am. Chem. Soc. 124:2394-5; Tang et al. (2006) Anal. Chem. 78:711-7 (2006)).
There is also an important need in the biomedical community for a reliable technology that produces the highest possible quality, reproducible antibody reagents to membrane proteins. Ongoing improvement of this technology pipeline will directly benefit both the health research community and the larger biomedical community.
The use of antibodies to detect single and multipass membrane bound proteins can be used to identify new biomarkers and perform many assays. For example, antibodies are also widely used in diagnostic applications, such as for clinical medicine (e.g., ELISA and radioimmunoassay systems). Analysis of cells and tissues in pathology laboratories includes the use of antibodies on tissue sections and in flow cytometry analyses. Antibodies are also useful as therapeutics.
The production of antibodies can be costly and time-consuming, thus methods for the high throughput production of antibodies, in particular, highly specific antibodies to membrane proteins, that is more cost-effective and less time-consuming is desirable. The present disclosure meets these needs, and provides related advantages.